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Mouse Sperm Cryopreservation and Recovery using the I·Cryo Kit
Published on: December 12, 2011
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Cryopreservation and Preparation of Thawed Spermatozoa from Rhesus Macaques (Macaca mulatta) for In Vitro
Fernanda M de Carvalho1, Cathy Ramsey2, Carol B Hanna2
1Department of Animal Reproduction, University of São Paulo, São Paulo, São Paulo, Brazil; Department of Reproductive & Developmental Sciences, Oregon National Primate Research Center, Oregon Health & Science University, Beaverton, Oregon;,
Summary
Optimizing sperm cryopreservation in rhesus macaques is crucial for disease modeling. Slow-freezing combined with density gradient centrifugation (DGC) yielded the most functional sperm for in vitro fertilization (IVF).
Area of Science:
- Reproductive Biology
- Animal Models
- Assisted Reproductive Technologies
Background:
- Rhesus macaques are vital models for human disease research, enhanced by gene editing and assisted reproductive technologies.
- Optimizing in vitro fertilization (IVF) in rhesus macaques is essential, as cryopreservation damage (cryodamage) negatively impacts success rates.
- Cryodamage to sperm can lead to failed artificial insemination and reduced fertilization and blastocyst formation.
Purpose of the Study:
- To systematically assess and compare two cryopreservation methods (vitrification and slow-freezing) and four sperm recovery techniques (simple wash, density gradient centrifugation, swim-up, glass wool filtration).
- To evaluate the efficacy of the optimal sperm recovery methods for in vitro fertilization (IVF) in rhesus macaques.
- To identify the most effective combination of cryopreservation and recovery methods for producing functional sperm for assisted reproduction.
Main Methods:
- Compared sperm survival rates after vitrification versus slow-freezing.
- Assessed four methods for removing cryoprotectants and isolating motile sperm post-thaw: simple wash (SW), density gradient centrifugation (DGC), swim-up (SU), and glass wool filtration (GWF).
- Evaluated the IVF success using sperm isolated by the two most effective recovery methods.
Main Results:
- Slow-freezing resulted in significantly higher sperm motility (42 ± 5%) compared to vitrification (1.2 ± 0.3%).
- Simple wash (SW) was superior for recovering total sperm and sperm with intact plasma membranes, but no method significantly differed in recovering motile sperm.
- IVF using sperm recovered by density gradient centrifugation (DGC) yielded 5% more embryos and 25% more blastocysts than using SW-recovered sperm.
Conclusions:
- Slow-freezing is a more effective cryopreservation method than vitrification for rhesus macaque sperm.
- Density gradient centrifugation (DGC) is the preferred method for isolating thawed sperm for IVF, leading to improved embryo and blastocyst development.
- The combination of slow-freezing and DGC offers the highest efficacy for producing functional rhesus macaque sperm for in vitro applications.
